Microbe-induced activation of inflammatory cytokine response in human cells

Human body is in continuous contact with microbes. Although many microbes are harmless or beneficial for humans, pathogenic microbes possess a threat to wellbeing. Antimicrobial protection is provided by the immune system, which can be functionally divided into two parts, namely innate and adaptive immunity. The key players of the innate immunity are phagocytic white blood cells such as neutrophils, monocytes, macrophages and dendritic cells (DCs), which constantly monitor the blood and peripheral tissues. These cells are armed for rapid activation upon microbial contact since they express a variety of microbe-recognizing receptors. Macrophages and DCs also act as antigen presenting cells (APCs) and play an important role in the development of adaptive immunity. The development of adaptive immunity requires intimate cooperation between APCs and T lymphocytes and results in microbe-specific immune responses. Moreover, adaptive immunity generates immunological memory, which rapidly and efficiently protects the host from reinfection. Properly functioning immune system requires efficient communication between cells. Cytokines are proteins, which mediate intercellular communication together with direct cell-cell contacts. Immune cells produce inflammatory cytokines rapidly following microbial contact. Inflammatory cytokines modulate the development of local immune response by binding to cell surface receptors, which results in the activation of intracellular signalling and modulates target cell gene expression. One class of inflammatory cytokines chemokines has a major role in regulating cellular traffic. Locally produced inflammatory chemokines guide the recruitment of effector cells to the site of inflammation during microbial infection. In this study two key questions were addressed. First, the ability of pathogenic and non-pathogenic Gram-positive bacteria to activate inflammatory cytokine and chemokine production in different human APCs was compared. In these studies macrophages and DCs were stimulated with pathogenic Steptococcus pyogenes or non-pathogenic Lactobacillus rhamnosus. The second aim of this thesis work was to analyze the role of pro-inflammatory cytokines in the regulation of microbe-induced chemokine production. In these studies bacteria-stimulated macrophages and influenza A virus-infected lung epithelial cells were used as model systems. The results of this study show that although macrophages and DCs share several common antimicrobial functions, these cells have significantly distinct responses against pathogenic and non-pathogenic Gram-positive bacteria. Macrophages were activated in a nearly similar fashion by pathogenic S. pyogenes and non-pathogenic L. rhamnosus. Both bacteria induced the production of similar core set of inflammatory chemokines consisting of several CC-class chemokines and CXCL8. These chemokines attract monocytes, neutrophils, dendritic cells and T cells. Thus, the results suggest that bacteria-activated macrophages efficiently recruit other effector cells to the site of inflammation. Moreover, macrophages seem to be activated by all bacteria irrespective of their pathogenicity. DCs, in contrast, were efficiently activated only by pathogenic S. pyogenes, which induced DC maturation and production of several inflammatory cytokines and chemokines. In contrast, L. rhamnosus-stimulated DCs matured only partially and, most importantly, these cells did not produce inflammatory cytokines or chemokines. L. rhamnosus-stimulated DCs had a phenotype of "semi-mature" DCs and this type of DCs have been suggested to enhance tolerogenic adaptive immune responses. Since DCs have an essential role in the development of adaptive immune response the results suggest that, in contrast to macrophages, DCs may be able to discriminate between pathogenic and non-pathogenic bacteria and thus mount appropriate inflammatory or tolerogenic adaptive immune response depending on the microbe in question. The results of this study also show that pro-inflammatory cytokines can contribute to microbe-induced chemokine production at multiple levels. S. pyogenes-induced type I interferon (IFN) was found to enhance the production of certain inflammatory chemokines in macrophages during bacterial stimulation. Thus, bacteria-induced chemokine production is regulated by direct (microbe-induced) and indirect (pro-inflammatory cytokine-induced) mechanisms during inflammation. In epithelial cells IFN- and tumor necrosis factor- (TNF-) were found to enhance the expression of PRRs and components of cellular signal transduction machinery. Pre-treatment of epithelial cells with these cytokines prior to virus infection resulted in markedly enhanced chemokine response compared to untreated cells. In conclusion, the results obtained from this study show that pro-inflammatory cytokines can enhance microbe-induced chemokine production during microbial infection by providing a positive feedback loop. In addition, pro-inflammatory cytokines can render normally low-responding cells to high chemokine producers via enhancement of microbial detection and signal transduction.Immuunijärjestelmä suojaa elimistöä taudinauheuttajamikrobeilta ja pahanlaatuisilta solumuutoksilta. Elimistön mikrobipuolustukseen osallistuvat seerumin antimikrobiset proteiinit ja erityisesti veren valkosolut. Veren ja kudosten neutrofiilit, monosyytit, makrofagit, sekä dendriittisolut tarkkailevat ympäristöään mikrobien ja muiden taudinaiheuttajien varalta. Nämä valkosolutyypit ilmentävät lukuisia mikrobireseptoreja, joiden avulla ne tunnistavat mikrobirakenteita, kuten bakteerien pinnan lipopolysakkarideja, peptidoglykaania tai virusten kaksijuosteista RNA:ta. Tunnistustapahtuma käynnistää solunsisäisen signaalinvälityksen, mikä johtaa valkosolujen geeniluennan aktivoitumiseen sekä toiminnallisiin muutoksiin. Valkosoluaktivaation tavoittena on pysäyttää infektion leviäminen mahdollisimman nopeasti. Solujen välinen viestintä on ehdoton edellytys toimivalle immuunijärjestelmälle. Liukoiset sytokiiniproteiinit ovat yksi tärkeimmistä immuunivasteen välittäjäaineista. Sytokiinigeenien luenta käynnistyy nopeasti mikrobikohtaamisen seurauksena. Nämä tulehdustyyppiset sytokiinit, eli pro-inflammatoriset sytokiinit, säätelevät immunivasteen kehittymistä mm. aktivoiden antimikrobisten tekijöiden tuotantoa. Eräs sytokiiniryhmä kemokiinit - säätelee valkosoluliikennettä tulehdusalueella. Tässä työssä on vertailtu patogeenisen Streptococcus pyogenes ja ei-patogeenisen Lactobacillus rhamnosus-bakteerin kykyä aktivoida ihmisen valkosoluja. Tulokset osoittavat, että S. pyogenes aktivoi inflammatorisen välittäjäainevasteen makrofageissa ja kaikissa tutkituissa dendriittisolutyypeissä. Ihmisen immuunivaste näyttää siis aktivoituvan tehokkaasti S. pyogenes-bakteeria vastaan, mikä osaltaan johtaa tämän bakteerin tuhoamiseen ja infektion leviämisen estämiseen. Toisaalta voimakas välittäjäainetuotanto ja siitä mahdollisesti seuraava ylisuuri immuunivasteen aktivaatio saattaa pahimmillaan johtaa kudostuhoon tai streptokokki-infektion jälkitauteina kehittyviin autoimmuunitauteihin. Ei-patogeeninen, elimistön normaaliflooraan kuuluva maitohappobakteeri L. rhamnosus aktivoi makrofagit ja dendriittisolut vain osittain eikä saanut aikaan inflammatoristen välittäjäaineiden tuotantoa dendriittisoluista. Inflammatorisen vasteen puuttuminen voi osaltaan selittää elimistön ja sen normaalimikrobiston välistä toleranssia ja symbioottista vuorovaikutusta. Tässä työssä on lisäksi selvitetty pro-inflammatoristen sytokiinien vaikutuksia solujen välittäjäaineiden tuotantoon bakteeri- ja virusinfektiossa. Tulokset osoittavat, että infektion aikana tuotettavat pro-inflammatoriset sytokiinit, erityisesti interferoni (IFN)-, lisäävät patogeenisen S. pyogenes-bakteerin aikaansaamaa välittäjäainetuotantoa, mikä edelleen tehostaa immuunivasteen aktivaatiota. IFN-:n ja tuumorinekroositekijä-:n osoitettiin myös lisäävän solujen mikrobitunnistusreseptorien ja signaalinvälityskomponenttien ilmentymistä, minkä seurauksena epiteelisolujen virusvaste tehostui merkittävästi. Tulokset osoittavat, että tehokas immuunivasteen aktivaatio riippuu sekä suorista mikrobi-kohdesolu vuorovaikutuksista että infektion aikana tuotettavien sytokiinien välittämistä signaaleista

Overcoming tumor resistance by heterologous adeno-poxvirus combination therapy

Successful cancer control relies on overcoming resistance to cell death and on activation of host antitumor immunity. Oncolytic viruses are particularly attractive in this regard, as they lyse infected tumor cells and trigger robust immune responses during the infection. However, repeated injections of the same virus promote antiviral rather than antitumor immunity and tumors may mount innate antiviral defenses to restrict oncolytic virus replication. In this article, we have explored if alternating the therapy virus could circumvent these problems. We demonstrate in two virus-resistant animal models a substantial delay in antiviral immune- and innate cellular response induction by alternating injections of two immunologically distinct oncolytic viruses, adenovirus, and vaccinia virus. Our results are in support of clinical development of heterologous adeno-/vaccinia virus therapy of cancer.Peer reviewe

Live Lactobacillus rhamnosus and Streptococcus pyogenes differentially regulate Toll-like

receptor (TLR) gene expression in human primary macrophage

Probiotic Leuconostoc mesenteroides ssp. cremoris and Streptococcus thermophilus induce IL-12 and IFN-γ production

AIM: To investigate the capacity of potentially probiotic strains from six bacterial genera to induce cytokine production alone or in combinations in order to identify potential enhancing or synergistic effects in order to select probiotic bacteria for in vivo purposes